Journal: bioRxiv

Article Title: Human cytomegalovirus virion delivered UL14 is a viral tropism factor for epithelial cells

doi: 10.64898/2026.05.18.725932

Figure Lengend Snippet: a ) Nuff-1 cells were infected at an MOI of 1.0 TCID 50 /cell using the TB40/E-mCherry-UL99eGFP virus. RNA was collected at 4 and 24 hours and UL14 transcripts were quantified by qPCR and normalized to GAPDH ( N = 3) b) Primary fibroblasts (Nuff-1) or c ) Epithelial cells (ARPE-19) were infected at a multiplicity of 3.0 TCID 50 /cell using the TB40/E-mCherry-UL99eGFP-UL14-FLAG virus. Cell lysates were collected across a time course of 96 hours and probed by western blot for the UL14 protein using anti-flag; β-Tubulin was used as the loading control. Representative blots are shown ( N = 3 ). d ) Primary fibroblasts were infected at an MOI of 1.0 TCID 50 /cell and treated with vehicle or PAA (300ug/mL) and harvested at 48 and 72 hours. Cell lysates were probed by western blot for UL14, pp65, IE1, and β-Tubulin. ( N = 3 )

Article Snippet: hTert MRC-5 embryonic lung fibroblast (passage 10-30), primary newborn human foreskin fibroblasts (NuFF-1; Global Stem, passage 10-30) and adult retinal pigmented epithelial cells (ARPE-19; ATCC; passage 34-40) were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM), supplemented with 10% fetal bovine serum (FBS), 2mM L-glutamine (523-100p, Cleveland Clinic Media Core), and 100 U/mL penicillin-streptomycin solution (721-100p, Cleveland Clinic Media Core).

Techniques: Infection, Virus, Western Blot, Control

Journal: bioRxiv

Article Title: Human cytomegalovirus virion delivered UL14 is a viral tropism factor for epithelial cells

doi: 10.64898/2026.05.18.725932

Figure Lengend Snippet: a) Primary fibroblasts (NuFF-1) were infected with the viruses indicated at an MOI of 1.0 TCID 50 /cell. Supernatants were collected over a time course of 120 h, and then viral titers were quantified for each time point by TCID 50 assays. b ) Epithelial cells (ARPE-19) were infected with the indicated viruses at a multiplicity of infection of 1.0 TCID 50 /cell. Cell associated virus was collected over a time course of 15 days, and then viral titers were quantified for each time point byTCID 50 assays. Mean ± SD is shown ( N = 3 ).

Article Snippet: hTert MRC-5 embryonic lung fibroblast (passage 10-30), primary newborn human foreskin fibroblasts (NuFF-1; Global Stem, passage 10-30) and adult retinal pigmented epithelial cells (ARPE-19; ATCC; passage 34-40) were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM), supplemented with 10% fetal bovine serum (FBS), 2mM L-glutamine (523-100p, Cleveland Clinic Media Core), and 100 U/mL penicillin-streptomycin solution (721-100p, Cleveland Clinic Media Core).

Techniques: Infection, Virus

Journal: bioRxiv

Article Title: Human cytomegalovirus virion delivered UL14 is a viral tropism factor for epithelial cells

doi: 10.64898/2026.05.18.725932

Figure Lengend Snippet: Epithelial cells (ARPE-19) were infected with the indicated viruses at a multiplicity of infection of 1.0 TCID 50 /cell. Images were taken at 3, 6, 9, and 12 days post infection with a BIO-RAD ZOE Fluorescent Cell Imager, where mCherry is being used as a marker of infection. Representative images are shown ( N = 3 ).

Article Snippet: hTert MRC-5 embryonic lung fibroblast (passage 10-30), primary newborn human foreskin fibroblasts (NuFF-1; Global Stem, passage 10-30) and adult retinal pigmented epithelial cells (ARPE-19; ATCC; passage 34-40) were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM), supplemented with 10% fetal bovine serum (FBS), 2mM L-glutamine (523-100p, Cleveland Clinic Media Core), and 100 U/mL penicillin-streptomycin solution (721-100p, Cleveland Clinic Media Core).

Techniques: Infection, Marker

Journal: bioRxiv

Article Title: Human cytomegalovirus virion delivered UL14 is a viral tropism factor for epithelial cells

doi: 10.64898/2026.05.18.725932

Figure Lengend Snippet: Epithelial cells (ARPE-19) were infected with the respective viruses at a multiplicity of infection of 3.0 TCID 50 /cell. a ) Cell lysates were collected at 48 hours post infection and IE1 was probed by western blot using the anti-iE1 antibody; β-Tubulin was used as the loading control. b ) Densitometry mean ± SD is shown, statistical significance calculated using t -test; *** p ≤ 0.0001 ( N = 3 ). c ) Viral glycoprotein and pp65 loading of cell-free virions was determined by western blot and pp150 served as a loading control ( N = 3 ). d ) Viral and cellular DNA were quantified by qPCR using primers targeting the UL123 region of the viral genome and MDM2 targeting the human genome, respectively. Genome copy numbers were determined through the employment of a BAC that contains both UL123 and MDM2, mean ± SD is shown ( N=3 ).

Article Snippet: hTert MRC-5 embryonic lung fibroblast (passage 10-30), primary newborn human foreskin fibroblasts (NuFF-1; Global Stem, passage 10-30) and adult retinal pigmented epithelial cells (ARPE-19; ATCC; passage 34-40) were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM), supplemented with 10% fetal bovine serum (FBS), 2mM L-glutamine (523-100p, Cleveland Clinic Media Core), and 100 U/mL penicillin-streptomycin solution (721-100p, Cleveland Clinic Media Core).

Techniques: Infection, Western Blot, Control

Journal: bioRxiv

Article Title: Human cytomegalovirus virion delivered UL14 is a viral tropism factor for epithelial cells

doi: 10.64898/2026.05.18.725932

Figure Lengend Snippet: a ) schematic of ΔUL14 virus passaged through UL14-FLAG stable expressing fibroblasts to generate UL14-Comp virus. b ) Fibroblasts were infected with the respective viruses at an MOI of 10 TCID 50 /cell and cell lysates were harvested at 6 hours post-infection and then used for western blotting, where UL14 was probed using anti-flag and β-Tubulin was used as a control ( N = 3 ). c ) fibroblasts or d ) epithelial cells were infected with the respective viruses at an MOI of 3.0 TCID 50 /cell. Cell lysates were collected at 48 hours post-infection and then used for western blotting. IE1 was probed for using anti-iE1 antibody and β-Tubulin was used as a loading control ( N = 3 ).

Article Snippet: hTert MRC-5 embryonic lung fibroblast (passage 10-30), primary newborn human foreskin fibroblasts (NuFF-1; Global Stem, passage 10-30) and adult retinal pigmented epithelial cells (ARPE-19; ATCC; passage 34-40) were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM), supplemented with 10% fetal bovine serum (FBS), 2mM L-glutamine (523-100p, Cleveland Clinic Media Core), and 100 U/mL penicillin-streptomycin solution (721-100p, Cleveland Clinic Media Core).

Techniques: Virus, Expressing, Infection, Western Blot, Control

Journal: bioRxiv

Article Title: Human cytomegalovirus virion delivered UL14 is a viral tropism factor for epithelial cells

doi: 10.64898/2026.05.18.725932

Figure Lengend Snippet: Epithelial cells were infected with the indicated viruses at an MOI of 5.0 TCID 50 /cell for 15 hours. Cells were fixed, permeabilized, and stained with EEA1 (Early Endosome Antigen I), pp65, and DAPI. Images were acquired using a 63x oil immersion objective. Scale bars 5 µm. ( N = 3); representative images are shown.

Article Snippet: hTert MRC-5 embryonic lung fibroblast (passage 10-30), primary newborn human foreskin fibroblasts (NuFF-1; Global Stem, passage 10-30) and adult retinal pigmented epithelial cells (ARPE-19; ATCC; passage 34-40) were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM), supplemented with 10% fetal bovine serum (FBS), 2mM L-glutamine (523-100p, Cleveland Clinic Media Core), and 100 U/mL penicillin-streptomycin solution (721-100p, Cleveland Clinic Media Core).

Techniques: Infection, Staining

Journal: bioRxiv

Article Title: Human cytomegalovirus virion delivered UL14 is a viral tropism factor for epithelial cells

doi: 10.64898/2026.05.18.725932

Figure Lengend Snippet: a ) Schematic of UL14 and UL141 viral mutants generated using BAC recombineering in the TB40/E background. b) Epithelial cells (ARPE-19) were infected with the indicated viruses at a multiplicity of infection of 1.0 TCID 50 /cell. Cell associated virus was collected over a time course of 15 days and then viral titers were quantified for each time point by TCID 50 assays. Mean ± SD is shown ( N=3 ).

Article Snippet: hTert MRC-5 embryonic lung fibroblast (passage 10-30), primary newborn human foreskin fibroblasts (NuFF-1; Global Stem, passage 10-30) and adult retinal pigmented epithelial cells (ARPE-19; ATCC; passage 34-40) were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM), supplemented with 10% fetal bovine serum (FBS), 2mM L-glutamine (523-100p, Cleveland Clinic Media Core), and 100 U/mL penicillin-streptomycin solution (721-100p, Cleveland Clinic Media Core).

Techniques: Generated, Infection, Virus