High G+C Content of Herpes Simplex Virus DNA: Proposed Role in Protection Against Retrotransposon Insertion

Jay C Brown*

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© Jay C. Brown; Licensee Bentham Open.

open-access license: This is an open access article licensed under the terms of the Creative Commons Attribution Non-Commercial License (http: //creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted, non-commercial use, distribution and reproduction in any medium, provided the work is properly cited.

* Address correspondence to this author at the Department of Microbiology Box 800734, University of Virginia Health System, 1300 Jefferson Park Avenue, Charlottesville, VA 22908, USA; Tel: 01-434 924 1814; Fax: 01-434 982 1071; E-mail: jcb2g@virginia.edu


The herpes simplex virus dsDNA genome is distinguished by an unusually high G+C nucleotide content. HSV-1 and HSV-2, for instance, have GC contents of 68% and 70% respectively, while that of the host (human) genome is 41%. To determine how GC content varies with genome location, GC content was measured separately in coding and intergenic regions of HSV-1 DNA. The results showed that the 75 genes constitute a uniform population with a mean GC content of 66.9 ± 4.1%. In contrast, intergenic regions were found in two non-overlapping populations, one with a mean GC content (69.3 ± 4.6% n=32) similar to the coding regions and another where the GC content is lower (56.0 ± 4.9 n=30). Compared to other regions of the genome, intergenic regions with reduced GC content were found to be enriched in local GC minima, CACACA sequences and a primary target sequence (TTAAAA) for retrotransposition events. The results are interpreted to suggest that a high GC content is part of the way HSV-1 protects its genes from invasion by mobile genetic elements active during cell differentiation in the nervous system.

Keywords: Herpes simplex virus, DNA sequence, G+C content, intergenic DNA, L1 retrotransposition, CA repeats.


The basic features of the herpes simplex virus (HSV-1) genome are now well known. Each virion contains a single molecule of dsDNA 152 kb in length. A total of 75 genes for known proteins are encoded with 69 of these present in a single copy and three in two copies each [1-3]. The genome is divided into two segments, L and S, which encode 57 (UL) and 12 (US) single copy genes, respectively. The L and S segments are separated by repeated genes, which are also found at the genome ends (Fig. 1). As HSV-1 DNA is replicated, the L and S segments invert with respect to each other yielding four genome “isomers” that are found in equal proportions in wild-type HSV-1 populations [4-6].

Fig. (1).

Schematic drawing of the HSV-1 genome. The genome is 152,261 bp in length and is divided into two segments, L and S.Each segment is organized with single copy genes (UL and US) in the middle and repeated genes (RL and RS) at the ends.

Among the genes encoded by HSV-1 are 43 ancestral or core genes present in all α-, β- and γ-herpesviruses [3, 7, 8]. All 43 are located in UL and most are conserved genes involved in vital virus functions such as entry of the virus into a host cell, DNA replication, capsid assembly, packaging DNA into the capsid and exit of the capsid from the host cell nucleus. The non-core HSV-1 genes include all US genes and highly divergent genes found at the segment ends [9]. Non-core genes encode proteins involved in lineage- or species-specific functions such as transcriptional transactivation, immune evasion and host cell recognition.

A high GC content is one of the most unusual features of the HSV-1 genome. HSV-1 and HSV-2 have GC contents of 68% and 70%, respectively, while the value for the host (human) genome is 41%. In contrast, 54.4 ± 11.5% is the average GC content for the 44 herpesviruses whose genome sequences have been determined (as of March 2007). Only eight herpesviruses, all α-herpesviruses, have GC contents of 68% or greater. In an effort to understand the reason for the high GC content, I have focused on HSV-1 and examined the extent to which GC content varies with position in the genome. The GC content was determined in the coding regions and in the regions between genes. The results have identified a group of intergenic regions whose GC content is reduced compared to the coding regions and to the genome as a whole.


The HSV-1 DNA sequence (NC_001806) was extracted from public databases and manipulated with programs in GCG (version 11.1.3; Accelrys) run on a Dell PowerEdge 1950 server with dual 3 GHz Xeon cpu’s and 10GB of RAM). Composition was used to determine GC contents and FindPatterns+ was used to locate specific sequences. GC content in a sliding 120 bp window was determined with Artemis (Sanger Institute). Local GC minima were identified by visual inspection of the trace with all identified minima at least 10% GC lower than the adjacent background.


GC Content in Coding and Intergenic Regions

Coding and intergenic regions were extracted from the HSV-1 DNA sequence (NC_001806) using the editor function of GCG, and GC contents were determined with Composition. The 75 genes were found to have a mean GC content of 66.9 ± 4.1% with a range of 58.1% (UL1) to 82.9% (RL1; Table 1). Gene GC contents constituted a single population centered at the mean value as shown in Fig. 2 (black bars). The GC contents of gene subgroups such as essential and non-essential genes or core and non-core genes were examined and found not to differ significantly from each other or from the genome as a whole (Table 2).

Table 1.

GC Content of Coding and Intergenic Regions in the HSV-1 Genome

Coding Region Intergenic Region (rightward) Coding Region Intergenic Region (rightward)
Gene Length (bp) GC Length (bp) %GCa Gene Length (bp) %GC Length (bp) %GC
RL1 747 82.9 1003 68.1 UL36 9495 71.3 170 60.6
RL2 (ex 2) 1604 77.2 5588 67.8 UL37 3373 69.3 448 68.3
UL1 675 58.1 Overlap UL38 1398 71.3 517 61.5
UL2 1005 66.1 70 48.6 UL39 3414 65.7 70 74.3
UL3 708 63.4 160 46.3 UL40 1023 61.4 221 53.4
UL4 600 64.8 62 69.4 UL41 1470 62.7 477 62.7
UL5 2649 62.1 Overlap UL42 1526 66.8 111 55.0
UL6 2031 68.3 Overlap UL43 1305 72.4 280 62.1
UL7 890 66.1 200 55.0 UL44 1516 67.8 187 57.8
UL8 2253 70.4 238 72.7 UL45 519 68.6 247 56.7
UL9 2555 63.5 Overlap UL46 2157 71.1 84 67.9
UL10 1445 65.3 154 53.2 UL47 2082 73.1 492 63.2
UL11 291 66.6 Overlap UL48 1473 65.1 370 64.3
UL12 1881 68.5 60 66.7 UL49 906 70.5 Overlap
UL13 1557 64.0 Overlap UL49A 1276 68.9 18 50.0
UL14 660 65.9 106 73.6 UL50 1116 66.7 153 48.4
UL15 (ex 1) 1029 64.3 127 60.6 UL51 735 68.4 38 76.5
UL16 1122 68.3 92 81.5 UL52 3177 66.1 Overlap
UL17 2112 70.1 139 66.2 UL53 1017 61.1 540 64.6
UL15 (ex 2) 1179 61.5 283 56.5 UL54 1539 69.3 225 60.0
UL18 957 65.5 354 68.1 UL55 561 62.4 166 50.6
UL19 4125 68.5 293 68.6 UL56 705 66.2 3766 68.3
UL20 669 61.3 587 60.3 RL2 1604 77.2 1003 68.1
UL21 1622 66.0 172 60.5 RL1 747 82.9 1375 77.3
UL22 2517 66.7 291 60.8 RS1 3897 81.4 1537 74.6
UL23 1304 63.5 Overlap US1 1297 64.9 94 61.7
UL24 1008 64.6 70 51.4 US2 876 64.3 295 65.1
UL25 1743 68.0 255 60.0 US3 1446 63.6 78 61.5
UL26 1908 71.4 343 47.8 US4 717 63.7 272 59.6
UL27 3023 66.5 9 77.7 US5 279 65.9 411 58.6
UL28 2358 69.7 305 66.6 US6 1185 64.3 183 60.6
UL29 3591 67.3 755 63.3 US7 1173 65.6 287 56.1
UL30 3708 65.8 Overlap US8 1653 66.5 419 69.2
UL31 921 65.6 Overlap US9 273 63.0 573 57.1
UL32 1791 68.0 Overlap US10 939 67.3 Overlap
UL33 393 67.9 81 70.4 US11 486 67.3 66 68.2
UL34 828 67.1 107 67.3 US12 267 65.5 1549 75.1
UL35 339 65.8 146 48.6 RS1 3897 81.4 1261 78.5

Intergenic regions with reduced GC content are highlighted. Others are in the genome-like group

Fig. (2).

Histogram showing the GC content of HSV-1 genes (black bars) and intergenic regions (gray bars). Note the presence of a single population of coding region GC contents. Intergenic GC contents are considered to fall into two distinct groups, one coinciding with the population of genomic GC contents and the other with lower GC content. The two groups are identified as genome-like and reduced GC content,respectively. The 32 intergenic regions in the genome-like group have a mean GC content of 69.3% (range 62.1%-81.5%) while in the reduced GC content group the mean is 56.0% (30 regions; range 46.3%-61.7%; indicated by the horizontal gray line). Not represented in this analysis are the regions between the left genome end and RL1, and between the right genome end and RS1.

Table 2.

Mean GC Content of Selected HSV-1 Gene Groupsa

Gene group Mean %GC ± SD P valueb
Essential 67.3 ± 3.9 (n=34) 0.55
Non-essential 66.5 ± 4.3 (n=38)
Core 66.7 ± 2.8 (n=43) 0.39
Non-core 67.3 ± 5.4 (n=38)
Beta kinetic class 65.5 ± 2.8 (n=13) 0.87
Gamma kinetic class 66.5 ± 4.3 (n=38)
All UL 66.6 ± 3.1 (n=58) 0.91
All US 65.2 ± 1.4 (n=12)

Source: Essential, Non-essential, Beta (early) kinetic class, Gamma (late) kinetic class: [7]; Core, Non-core: [3]; UL, US: GenBank NC_001806.

Applies to the hypothesis that the GC content of the two gene groups is different.

Compared to gene GC contents, intergenic regions were found to have a broader range of values, which extend from 46.3% (UL3-UL4) to 81.5% (UL16-UL17). Intergenic region GC contents were considered to form two non-overlapping populations as shown in Fig. 2 (gray bars). One coincided approximately with the distribution of gene GC contents while the other spanned a range of lower values. Further analysis was done with the two populations, which are identified as the genome-like GC content group (32 regions) and reduced GC group (30 regions), respectively (Table 1). Intergenic regions in the genome-like group had GC contents in the range of 62.1% (UL43)-81.5% (UL16); the range of the reduced GC group was 46.3% (UL3)-61.7% (US1).

Both genome-like and reduced GC intergenic regions are found throughout the HSV-1 genome (Table 1). In each case, however, there are areas of concentration. Reduced GC regions, for instance, occur preferentially at three sites, UL20-UL26, UL35-UL45 and US3-US7 that contain evolutionarily divergent genes (Table 1) [9]. In contrast, genome-like intergenic regions are enriched in two areas, UL16-UL20 and UL27-UL34, where evolutionarily conserved genes are concentrated (Table 1).Genome-like intergenic regions are also enriched between genes found near the genome ends and near the inter-segment junction.

The lengths of intergenic sequences vary from zero (i.e. overlapping genes) to several hundred base pairs (excepting very long intergenic regions that occur near the segment ends). Genome-like and reduced GC intergenic regions differ only slightly in average length, the values being 258.7±195.1 and 230±138.8, respectively. Many reduced GC intergene lengths occur in a single population with a mean of approximately 150 bp, while genome-like lengths are more evenly distributed in the range of 0-600 bp (Fig. 3).

Fig. (3).

Histogram showing the lengths of HSV-1 intergenic regions with genome-like GC content (top) and reduced GC content (bottom). Note the population of reduced GC intergenic lengths centered at ~150 bp. Excluded from this analysis are very long intergenic regions between RL1-RL2, RL2 (exon 2)-UL1, UL56-RL2, RS1-US1 and US12-RS1.

Local GC Minima, CA Repeats and TTAAAA Sites

Inspection of intergenic sequences revealed additional features that distinguish genome-like from reduced GC regions. For instance, measurement of GC content in a sliding window 120 bp in length showed that intergenic regions were often the sites of pronounced local minima in GC content. This feature is illustrated in Fig. 4, which shows the GC content in two regions of the genome. Local GC minima were found to occur, for instance, between UL42-UL43, UL43-UL44, and US5-US6, but not between US6-US7. The presence of local GC minima is indicated for intergenic regions with genome-like and reduced GC content in Tables 3 and 4, respectively. Of the 29 intergenic regions with reduced GC content, 19 were found to be sites of local GC minima while only 8 of 29 genome-like intergenic regions have the same feature.

Fig. (4).

GC content as measured in a sliding 120 bp window shown in representative regions of the HSV-1 genome (between UL42-UL46 and US5-US10). Note the presence of pronounced local minima in GC content in certain intergenic regions such as UL42-UL43 and UL43-UL44, but not in others such as US6-US7.

Table 3.

Intergenic Regions with Genome-Like GC Contenta

Intergenic Region Local GC minb CACACA TTAAAA
RL1-RL2 No No Yes (2)
RL2 (ex 2)-UL1 Yes Yes (2) No
UL4-UL5 No No No
UL8-UL9 No No No
UL12-UL13 No No No
UL14-UL15 (ex 1) No No No
UL16-UL17 No No No
UL17-UL15 (ex 2) No No No
UL18-UL19 No No No
UL19-UL20 No Yes No
UL28-UL29 No Yes No
UL29-UL30 Yes No No
UL33-UL34 No No No
UL34-UL35 No Yes No
UL37-UL38 No No No
UL39-UL40 No No No
UL41-UL42 Yes No No
UL43-UL44 Yes Yes No
UL46-UL47 No No No
UL47-UL48 Yes Yes No
UL48-UL49 No No No
UL51-UL52 No No No
UL53-UL54 No No No
UL56-RL2 Yes No No
RS1-US1 Yes No No
US2-US3 No No Yes
US8-US9 No Yes Yes
US11-US12 No No No
US12-RS1 Yes No No
Total 8 8 4
Total per 10,000 bp 4.1 4.1 2.0

Intergenic regions with genome-like GC content are those with GC contents in the range of 62.1%-81.5%

Local GC minima are those identified by visual inspection of the GC content trace as defined in a sliding 120 bp window as shown in Fig. ((4)).

Table 4.

Intergenic Regions with Reduced GC Contentaa

Intergenic Region Local GC Minb CACACA TTAAAA
UL2-UL3 Yes No No
UL3-UL4 Yes No No
UL7-UL8 No Yes No
UL10-UL11 Yes Yes Yes (2)
UL15 (ex 1)-UL16 >No Yes (2) No
UL15 (ex 2)-UL18 Yes Yes No
UL20-UL21 Yes No No
UL21-UL22 No No No
UL22-UL23 Yes No No
UL24-UL25 Yes No No
UL25-UL26 No No No
UL26-UL27 Yes Yes (3) Yes
UL35-UL36 Yes No Yes
UL36-UL37 No No No
UL38-UL39 Yes Yes No
UL40-UL41 No No No
UL42-UL43 Yes Yes (2) No
UL44-UL45 Yes No Yes
UL45-UL46 Yes Yes No
UL50-UL51 Yes Yes No
UL54-UL55 No No No
UL55-UL56 Yes Yes No
US1-US2 Yes No No
US3-US4 No No No
US4-US5 No Yes No
US5-US6 Yes Yes Yes
US6-US7 No No No
US7-US8 Yes No No
US9-US10 Yes No No
Total 19 16 6
Total per 10,000 bp 28.4 23.9 9.0

Intergenic regions with reduced GC content are those with GC contents in the range of 46.3%-61.7%.

Local GC minima are those identified by visual inspection of the GC content trace as defined in a sliding 120 bp window as shown in Fig. ((4)).

The HSV-1 genome was examined for the frequency of two specific sequence motifs, CA repeats and TTAAAA sites. CA repeats are a common feature of the host (human) genome where they constitute 0.3% of the total sequence [10]. CA repeats occur predominantly as microsatellite DNA, but other functions have also been defined (see below). In the HSV-1 genome the sequence CACACA was taken as an overall measure of the CA repeat frequency. It occurs 79 times, a value in excess of the statistical expectation for a random 68% GC genome (49 see Table 5). Compared to the statistical expectation, CACACA sequences are enriched in intergenic regions; 17 are expected on a statistical basis and 24 are found (Table 5). Further, among the 24 intergenic CACACA sequences, most (16) are found in reduced GC regions (Tables 4 and 5).

Table 5.

CA Repeat and TTAAAA Retrotransposition Insertion Sites in the HSV-1 Genomea

Sequences Expecteda Observed
CACACA (total) 49 79
CACACA (in genes) 39 55
CACACA (intergenic regions) 10 24
TTAAAA (total) 5 28
TTAAAA (in genes) 4 18
TTAAAA (intergenic regions) 1 10

Statistically expected sequence numbers were calculated based on both strands of the 152,261 bp HSV-1 genome with 68% GC content. A 16%; T 16%; G 34%; C 34%. Calculated values were rounded to the nearest whole number. The proportions of gene and intergene regions in the HSV-1 genome were taken as 79% and 21%, respectively [1, 2]

TTAAAA sequences were identified because these are prominent target sites for introduction of novel DNA by retrotransposition [11-14]. Such DNA inserts, including the long interspersed nuclear elements (LINE or L1 sequences) could affect the virus by inactivating it if incorporation was into a required gene. Retrotransposition could also affect the genome by introduction of a novel gene into the HSV-1 genome if incorporation follows reverse transcription of a cellular mRNA [15-18].

Analysis demonstrated that the HSV-1 genome contains a total of 28 TTAAAA sequences, a value greater than the number expected on a statistical basis (Table 5). Eighteen occur in coding regions and 10 in intergenic segments. Of those in intergenic regions, 6 are found in reduced GC and 4 in genome-like sequences (Tables 3 and 4. Of the four present in genome-like intergenic regions, all are in intergenic segments (at the ends of UL and in US) that contain evolutionarily divergent genes [9].


Intergenic Regions with Reduced GC Content

The high GC content observed for HSV-1 genes was fully expected in light of the high GC content of the genome as a whole and of the RL, RS, UL and US regions [2]. It was remarkable to notice, however, that coding region GC contents occur in a single, relatively narrow distribution (Fig. 2). In view of the diversity of HSV-1 genes, it might have been expected that different populations would be observed corresponding, for instance, to essential compared to non-essential genes, UL compared to US genes, core compared to lineage specific genes or early (beta) compared to late (gamma) genes (see Table 2). The existence of a single population indicates that coding region GC content is affected by factors that apply to all genes in a more or less equal way.

The greater diversity of GC contents in intergenic regions contrasts with the narrow distribution in the genes (Fig. 2). While some intergenic GC contents are similar to the genes and to the genome as a whole, there are others with lower GC contents. These are distributed over a wide range (~46%- ~61%) as shown in Table 1 and Fig. 2. The presence of a reduced GC population of intergenic regions suggests that evolutionary forces constraining other sequences apply differently in this case, and understanding the difference might be revealing about HSV-1 evolution, replication and pathogenesis.

High GC content in genes and the effects of L1 retrotransposition

I suggest the high GC content of genes may be related to the way HSV-1 adapts to retrotransposition events in neurons. HSV-1 enters the host by infecting epithelial cells, but it promptly traffics to neurons in local (trigeminal) ganglia where it establishes a latent infection that most often lasts for the lifetime of the host. The virus can be reactivated from neurons causing recurrent infections that are the hallmark of HSV-1 disease [19].

Neurons arise during development from progenitor cells that undergo a large-scale proliferation accompanied by generation of extensive cell diversity [20, 21]. Functioning neurons are selected from this highly diverse population that may initially include 1010 or more cell types. Selection is based on the activity of individual neurons. Those that become incorporated into active networks survive while the others are lost. It is estimated that no more than 15%-40% of post-mitotic neurons survive this experience-based selection process [22-23].

Although the mechanisms used for generation of neuronal cell diversity are not thoroughly understood, it is considered that retrotransposition by L1 elements plays a significant role [23]. The human genome contains approximately 105 L1 elements (non-LTR retrotransposons) that together account for ~15% of the genome. Active L1 elements are 5 kb-6 kb in length, but most L1s are inactive due to 5’ deletions, rearrangements or mutations in the open reading frames. Only 80-100 human L1s are considered to be retrotransposition competent, and of these ~10% are considered to be highly active [11, 16].

Two genes, an endonuclease and reverse transcriptase are encoded in L1 retrotransposons, and transposition involves reverse transcription of the L1 mRNA. Reverse transcription takes place at specific sites in the host cell genome where primers are created following single-strand cuts introduced by the L1-encoded endonuclease. L1 elements are introduced at these sites, which occur preferentially at TTAAAA and related AT-rich sequences [11-14]. As a rare event, the L1 reverse transcription machinery can act in trans on cellular mRNAs to introduce genes and processed pseudogenes at L1 retrotransposition sites [17, 18].

In the developing nervous system, L1 retrotransposition can, in principle, contribute to generation of cell diversity by any process in which insertion of exogenous DNA can affect gene expression. Introduction into an exon, for instance, could block protein expression while similar insertion events could affect promoter function, alternative splicing or other processes. Recent studies have demonstrated that L1 retrotransposition can promote other forms of genetic instability by way of the DNA repair machinery [11]. L1 retrotransposition has been observed experimentally in mice, in HeLa cells and in neural progenitor cells in culture [11, 24]. In the latter case, retrotransposition is found to be enhanced following downregulation of Sox2, a presumptive inhibitor of L1 mobility [24].

It is suggested that a high GC content is a part of the way HSV-1 protects itself from harmful effects that might result from L1 insertion, particularly insertion into genes. A high GC content is expected to minimize the number of TTAAAA and other AT-rich insertion sites present in genes and therefore to protect the genes from L1 retrotransposition events. Such protection would be particularly beneficial in the case of latently infected cells where the number of HSV-1 genomes is reduced compared to lytic infections [25]. The proposed high rate of L1 retrotransposition in neurons makes a high GC content particularly attractive for viruses such as HSV-1 that replicate and establish latent infections in neurons. It is consistent with this idea to note that of the 44 sequenced herpesviruses, all eight with GC contents of 68% or greater have a tropism for the nervous system. (The eight are: HSV-1, 68% HSV-2, 70% cercopithecine herpesvirus 1, 74% cercopithecine herpesvirus 2, 70% cercopithecine herpesvirus 16, 76% bovine herpesvirus 1 72% bovine herpesvirus 5, 74% pseudorabies virus, 73%.) In contrast, varicella-zoster virus (VZV) has a comparatively low GC content (46%) despite the fact that it becomes latent in neurons [26]. The ability of VZV to produce such infections despite a low GC content may be due at least in part to the diversity of ganglia in which latent infections occur. Latent VZV infections are found in ganglia along the entire length of the central neural axis [26] while those of HSV-1 and HSV-2 are concentrated in a more restricted number of ganglia (e.g. the trigeminal in the case of HSV-1).

Retrotransposition and Reduced GC Intergenic Regions

The existence of a population of intergenic regions with reduced GC content may also be related to retrotransposition events in neurons. Among the ways herpesviruses are thought to adapt to environmental change, on an evolutionary time scale, is by incorporating entire genes from the host cell genome. Evidence for such incorporations has been obtained most recently in the case of certain gammaherpesviruses [3, 27-29]. Because the incorporated genes lack introns, it is presumed that their incorporation occurs by way of a cDNA intermediate and L1 retrotransposition. In the case of HSV-1, the existence of intergenic regions with reduced GC content would be preferred as sites for gene insertion due to the greater probability that they will contain AT-rich L1 insertion sequences. Such regions would arise by default if their creation involved simply resisting the evolutionary pressure favoring increased GC content. Evidence for preference of TTAAAA sequences in reduced GC compared to genome-like intergenic regions is described in this study (Tables 4 and 5).Six TTAAAA sequences are found in 6688 bp of reduced GC intergenic DNA while 4 are found in 19,651 bp of genome-like intergenic sequence.

Experimental Tests

In the future it should be possible to test the idea expressed here that a high GC content is involved in resistance of the HSV-1 genome to invasion by L1 retrotransposons. As noted above, it is now possible to observe L1 transposition in cultured cells with the L1 element derived either from the cell genome or from a transfected plasmid [11, 24, 30]. Similar tests with herpesvirus-infected cells should demonstrate that L1 transposition into the virus genome occurs at a higher rate in low GC compared to high GC viruses. In HSV-1-infected cells, transposition should occur preferentially into reduced GC intergene regions and should avoid genes.

Local GC Minima and CA Repeats

Determination of the HSV-1 GC content in a 120 bp window was carried out as a supplement to measurements of the gene and intergene values. It was expected that intergenic regions might have a reduced GC content because of the measurements described above and also because intergenic regions are often the sites of AT-rich polyadenylation signals (AATAAA). In spite of such expectations, it was nevertheless noteworthy that GC minima were as pronounced and as focused on intergenic regions as the results demonstrated (Fig. 4). Intergenic regions with pronounced GC minima were found throughout the HSV-1 genome with a preference for reduced GC intergenic sequences as shown in Tables 3 and 4. Further information about the function of intergenic GC minima might best be pursued by examining the genomes of other viruses for this feature, and appropriate studies are now underway in our laboratory.

CA dinucleotide repeats are an abundant feature of the human genome where they account for ~0.3% of the total sequence [10, 31]. Most occur in microsatellite sequences, but others are involved in functions such as regulation of gene expression [32, 33], control of alternative mRNA splicing [34, 35] and mRNA stability [36]. Such functions depend on the ability of CA repeats to bind heterogeneous nuclear ribonucleoprotein L (hnRNP L [33, 36]). In the HSV-1 genome, one CACACA sequence has been identified within the thymidine kinase coding region (UL23) where it acts by way of hnRNP L binding to enhance mRNA polyadenylation and nuclear export of intronless mRNAs [36]. Others of the 55 coding region CACACA sequences may act in the same or a similar way. CACACA sequences are also found in intergenic DNA. These are located throughout the HSV-1 genome with most (16 of 24) occurring in reduced GC intergenic regions (Tables 3 and 4). Analysis of deletions in such CACACA sequences suggests itself as a promising way to identify their function.


For contributions during the course of this investigation I gratefully acknowledge Michael Black, Fred Homa, Bill Newcomb Ann Campbell, Anna Maria Copeland, Lou Hammarskjöld and Jeffrey Brown. This work was supported by NIH award AI41466.


[1] McGeoch DJ, Dolan A, Donald S, Brauer DH. Nucleic Acids Res 1986; 14: 1727-45.
[2] McGeoch DJ, Dalrymple MA, Davison AJ, et al. J Gen Virol 1988; 69: 1531-74.
[3] McGeoch DJ, Rixon FJ, Davison AJ. Virus Res 2006; 117: 90-104.
[4] Roizman B, Knipe DM. Herpes simplex viruses and their replication In: Knipe DM, Howley PM, Eds. Fields Virology. 4th ed. Philadelphia: Lippincott Williams and Wilkins 2001; Vol 2: pp. 2399-459.
[5] Delius H, Clements JB. J Gen Virol 1976; 33: 125-33.
[6] Hayward GS, Jacob RJ, Wadsworth SC, Roizman B. Proc Natl Acad Sci USA 1975; 72: 4243-7.
[7] Roizman B. Proc Natl Acad Sci USA 1996; 93: 11307-12.
[8] Davison AJ. Vet Microbiol 2002; 86: 69-88.
[9] Brown J. Virology 2004; 330: 209-20.
[10] Lander ES, Linton LM, Birren B, et al. Nature 2001; 409(6822): 860-921.
[11] Gilbert N, Lutz S, Morrish TA, Moran JV. Mol Cell Biol 2005; 25: 7780-95.
[12] Cost GJ, Boeke JD. Biochem 1998; 37: 18081-93.
[13] Feng Q, Moran JV, Kazazian HH Jr., Boeke JD. Cell 1996; 87: 905-16.
[14] Jurka J. Proc Natl Acad Sci USA 1997; 94: 1872-7.
[15] Kazazian HHJr. Science 2004; 303: 1626-32.
[16] Kazazian HHJr, Moran JV. Nat Genet 1998; 19: 19-24.
[17] Esnault C, Maestre J, Heidmann T. Nat Genet 2000; 24: 363-7.
[18] Pavlicek A, Paces J, Elleder D, Hejnar J. Genome Res 2002; 12: 391.
[19] Whitley RJ, Knipe DM, Howley PM. Herpes simplex viuses In: Fields Virology. 4th ed. Philadelphia: Lippincott Williams and Wilkins 2001; Vol. 2: pp. 2461-509.
[20] Finlay BL, Slattery M. Science 1983; 219: 1349-51.
[21] Ferrer I, Soriano E, Del Rio JA, Alcantara S, Auladell C. Prog Neurobiol 1992; 39: 1-43.
[22] Oppenheim RW. Annu Rev Neurosci 1991; 14: 453-501.
[23] Muotri AR, Gage FH. Nature 2006; 441: 1087-93.
[24] Muotri AR, Chu VT, Marchetto MC, Deng W, Moran JV, Gage FH. Nature 2005; 435: 903-10.
[25] Wagner EK, Bloom DC. Clin Microbiol Rev 1997; 10: 419-43.
[26] Gilden DH, Mahalingam R, Cohrs RJ, Tyler KL. Nat Clin Pract Neurol 2007; 3: 82-94.
[27] Markine-Goriaynoff N, Georgin JP, Goltz M, et al. J Virol 2003; 77: 1784-92.
[28] Rivailler P, Jiang H, Cho YG, Quink C, Wang F. J Virol 2002; 76: 421-6.
[29] McGeoch DJ, Davison AJ. Semin Cancer Biol 1999; 9: 201-9.
[30] Ostertag EM, Prak ET, DeBerardinis RJ, Moran JV, Kazazian HHJr. Nucleic Acids Res 2000; 28: 1418-23.
[31] Venter JC, Adams MD, Myers EW, et al. Science 2001; 291(5507): 1304-51.
[32] Huang TS, Lee CC, Chang AC, et al. Biochem Biophys Res Commun 2003; 300: 901-7.
[33] Hui J, Reither G, Bindereif A. RNA 2003; 9: 931-6.
[34] Hui J, Hung LH, Heiner M, et al. EMBO J 2005; 24: 1988-98.
[35] Lee JH, Jeon MH, Seo YJ, et al. J Biol Chem 2004; 279(41): 42758-64.
[36] Guang S, Felthauser AM, Mertz JE. Mol Cell Biol 2005; 25(15): 6303-13.